KC-6097

293T-Rhesus-PTH1R Cell Line

62485

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Background of 293T-Rhesus-PTH1R Cell Line

PTH1R, also known as PTHR1, is a protein that belongs to the G-protein coupled receptor 2 family. PTH1R is a receptor for parathyroid hormone and for parathyroid hormone-related peptide. The activity of this receptor is mediated by G proteins which activate adenylyl cyclase, and phosphatidylinositol-calcium second messenger system. Diseases associated with PTH1R include Jansen type metaphyseal chondrodysplasia and Blomstrand type Chondrodysplasia. Among its related pathways are endochondral ossification with skeletal dysplasias and GPCR downstream signaling.

Specifications

Catalog Number	KC-6097
Cell Line Name	293T-Rhesus-PTH1R Cell Line
Clone Number	1#
Host Cell Line	293T
Description	Stable 293T cell line expressing exogenous human Rhesus-PTH1R gene
Quantity	One vial of frozen cells (≥2-10⁶/vial)
Stability	Stable in culture over a minimum of 10 passages
Application	Drug screening and biological assays
Freezing Medium	70% basal medium+20% FBS+10% DMSO
Propagation Medium	DMEM+10% FBS +1μg/mL Puromycin
Selection Marker	Puromycin
Morphology	Epithelial
Subculture	Split saturated culture 1:4 to 1:8 every 2-3 days; seed out at about 1-3 × 10⁵ cells/mL
Incubation	37 °C with 5% CO₂
Storage	Liquid nitrogen immediately upon receiving
Doubling Time	Approximately 30 hours
Mycoplasma Status	Negative
In Vivo Validation	NA

Cell Line Generation

293T-Rhesus-PTH1R cell line was generated using a lentiviral vector expressing the rhesus PTH1R sequence.

Characterization

Figure 1. Characterization of rhesus PTH1R overexpression in the 293T-rhesus-PTH1R stable clone using FACS.

Figure 2: Characterization of rhesus PTH1R in the 293T-rhesus-PTH1R stable clone using PCR sequencing.

Cell Resuscitation

Pre-warm complete culture medium (basal medium and 10% FBS) in a 37°C water bath.
Rapidly thaw the cryovial in a 37°C water bath for 1-2 minutes with gentle agitation.
Transfer the vial to a biosafety cabinet, and disinfect the exterior with 70% ethanol.
Aseptically transfer the cell suspension dropwise into a sterile centrifuge tube containing 9.0 mL of pre-warmed complete medium.
Centrifuge at approximately 125 × g for 5–7 minutes at room temperature, carefully aspirate the supernatant without disturbing the cell pellet.
Gently resuspend the pellet in an appropriate volume of complete medium and transfer the suspension into a T25 flask.
Incubate the flask in a 37°C in a humidified 5% CO₂ incubator.
Assess cell viability and morphology after 24 hours. If cells appear healthy, replace the medium with fresh medium supplemented with the appropriate selective antibiotic.
Subculture the cells at a ratio of 1:4 to 1:8 every 2-3 days upon reaching 80%–90% confluency.

Cell Freezing

Prepare the freezing medium (70% basal medium, 20% FBS and 10% DMSO) freshly before use.
Pre-chill the freezing medium on ice and label the cryovials accordingly.
Transfer the cell suspension to a sterile conical tube and perform a cell count to determine total viability and density.
Centrifuge the cells at 250×g for 5 minutes at room temperature; carefully aspirate the supernatant.
Gently resuspend the cell pellet in chilled freezing medium, ensuring a minimum cell density of 3×10⁶ cells/mL.
Aliquot 1 mL of the cell suspension into each pre-labeled cryovial.
Place the cryovials into a CoolCell® container and store at -80°C overnight for controlled-rate cooling.
Transfer the cryovials to the liquid nitrogen for long-term storage the following day.

References

Structure of the parathyroid hormone receptor C terminus bound to the G-protein dimer Gbeta1gamma2. PMID: 18611381 PMCID: PMC2601695 DOI: 10.1016/j.str.2008.04.010.
Dimeric arrangement of the parathyroid hormone receptor and a structural mechanism for ligand-induced dissociation. PMID: 20172855 PMCID: PMC2852981 DOI: 10.1074/jbc.M109.093138.
Yamaguchi T, Hosomichi K, Narita A, Shirota T, Tomoyasu Y, Maki K, Inoue I (Jul 2011). ""Exome resequencing combined with linkage analysis identifies novel PTH1R variants in primary failure of tooth eruption in Japanese"". Journal of Bone and Mineral Research. 26 (7): 1655–61. doi:10.1002/jbmr.385. PMID 21404329. S2CID 23855913.

Use License Agreement

Research Use Only.

Not for use in diagnostic procedures or therapeutic applications.

Redistribution of the cell line or its derivatives is prohibited without prior written permission from Kyinno Biotechnology.